High Schistosoma mansoni Co-Infection in Tuberculosis Patients with or without Human Immunodeficiency Virus: A Prospective Cohort Study

Background People with Latent tuberculosis infection (LTBI) remain the reservoir of tuberculosis. One-third to 1/4 of the world’s population is infected. Its reactivation is due to factors that disrupt the host’s immune response. Recent findings showed that Schistosoma mansoni coinfection leads to a Th2/Th1 profile which results in an immune modulation that favors the escape of the Mycobacteria. Schistosoma mansoni may contribute to TB incidence in endemic regions. We aimed to investigate the co-infection rate and patient outcomes. Methods A prospective cohort study was conducted between 2020–2022 at University Clinical Research Center (UCRC), including culture-confirmed active pulmonary TB patients and tested for Schistosoma mansoni in stools using Kato-Katz Technique. After descriptive analysis a logistic regression was performed to determine risk factors associated with TB and Schistosoma mansoni co-infection. Results Data of 174 tuberculosis-confirmed patients, Kato-Katz tested were analyzed. Males represented 62.6%, mean age was 34.9 ± 13.8 years, 29.9% were smokers, alcohol consumption 13.8%, TB contact history 26.4%, HIV coinfection 11.5%, diabetes 6.3%, undernourished 55.7%. Schistosoma mansoni prevalence was 28.7%. The co-infection was associated with less lung cavitation [aOR = 0.24 [95% CI (0.06–0.85), p = 0.028], unfavorable treatment result [aOR = 2.95 (1.23–7.08), p = 0.015] and death [aOR = 3.43 (1.12–10.58), p = 0.032]. Conclusions Despite Kato-Katz’s low sensitivity, Schistosoma mansoni coinfection was found in one-third of the TB patients; 2.5-fold higher than that of HIV. The coinfection was associated with poor treatment results and death.


Background
Tuberculosis (TB) was the leading cause of death worldwide due to a single infectious agent before the COVID-19 pandemic.In 2022, 10.6 million people became ill with TB of whom 1.3 million (15%) died (~ 4384 per day) [1].For the same year, TB killed more than HIV/AIDS by 2.5-fold (650,000) and malaria by 2.6-fold (619,000) [1,2].The new goals of the World Health Organization (WHO) to end TB are to reduce its mortality and incidence by 95% and 90%, respectively by 2035 [3].Latent tuberculosis infection (LTBI) is de ned as a state of persistent immune response to stimulation by Mycobacterium tuberculosis (Mtb) antigens without evidence of clinical manifestations of active TB.About one-third of the world's population and lives with the bacteria [4,5].LTBI is controlled by a T-helper 1 (Th1) in ammatory reaction that results in granulomas formation to inactive the bacteria, involving several immune cells such as macrophages, dendritic cells, lymphocytes, and cytokines including interleukins (IL)-12, 17, 23 and interferon-gamma (INF-).Despite the granulomas, 5-10% of people with LTBI will reactivate to active TB during their lifetime.Reactivations are due to phenomena that induce immune depression such as malnutrition, HIV/AIDS, cancer, diabetes, smoking, alcohol, etc. [4,5].Despite improved TB diagnostic tools and treatment strategies, the incidence remains high in low-and middle-income countries, particularly in Africa, Southern East Asia, and the Western Paci c. HIV/AIDS is one of the main risk factors of LTBI reactivation increasing the risk by 20 to 30-fold due to depletion of lymphocyte TCD4 + cells [6].Anti-tumor Necrosis Factor (anti-TNF) used in the treatment of systemic diseases also increases the risk of reactivation [7].In Canada, smokers had a 1.8-fold increased risk of LTBI and a 2.3fold risk of reactivation [8].Recent studies brought immunological evidence that Schistosoma mansoni (S. mansoni) co-infection with LTBI promotes the reactivation of TB bacteria by modulating the type Th1 immune response.During S. mansoni chronic infection, eggs disseminate throughout different organs and induce a type Th2 in ammatory reaction with the formation of granulomas against the eggs.Unlike, the type Th1 in LTBI, there is a proliferation of eosinophil cells and cytokines IL-4, 5, 10,13, and immunoglobulin E (IgE) [9].The coexistence of the type Th2 immune response modulates the type Th1, thus, promoting the escape and multiplication of the bacilli towards active TB [10][11][12].Pulmonary locations of S. mansoni eggs have been reported to represent 20-40% in the literature [13].Mali is one of the sub-Saharan African countries where the prevalence of S. mansoni is between 10 and 45% [14].A systematic review included two cross-sectional studies in Mali found a prevalence of 12.7% and 17.3% [15].To achieve WHO end TB goals, emphasis must be placed on the identi cation and management of unknown LTBI reactivation risk factors to reduce TB incidence and break transmission.Each risk factor must be considered in the context of epidemiology and endemicity.A better understanding of the impact of S. mansoni coinfection on LTBI control will help to design strategies of control to limit the number of LTBI reactivations in regions endemic to both infections.Thus, this study aimed to investigate the coinfection of S. mansoni in active pulmonary TB patients in Mali.We hypothesized that S. mansoni infection is higher among TB patients than HIV and the coinfection is associated with poor treatment outcomes.

METHODS
Study Design and Setting.This was a prospective cohort study over 30 months from January 1, 2021, to Study Population, Inclusion, and Exclusion criteria.All microscopy-positive pulmonary TB patients starting TB treatment were considered for this study.We included both males and females, aged ≥ 12 years, con rmed rifampicin sensitive by GeneXpert/RIF® and culture, naïve to TB treatment, permanent residents in Mali for > 5 years, who consented (≥ 18 years) or assented (12-17 years).We excluded patients with sputum cultures negative for Mycobacterium tuberculosis (Mtb) or positive for nontuberculous mycobacteria or rifampicin-resistant and/or multidrug-resistant tuberculosis.
Data and sample collection.At the day0 (D0) visit, a questionnaire was used for socio-demographic parameters such as age, sex, profession, permanent residence, living near river; medical histories such as history of TB contact, HIV, diabetes, and cancer; clinical parameters such as symptoms, BMI, blood pressure, and heart rate.Two sputum samples on 2 consecutive days (D0 and D1) and 5 mL blood sample were obtained.Chest x-ray results were collected, and ndings were classi ed into unilateral in ltrate, bilateral in ltrate, and the presence of a cavity.The treatment was monitored for 6 months to record outcomes.The regimen used in Mali is 2 months of Rifampicin (R), Isoniazid (H), Pyrazinamide (Z), and Ethambutol (E) and 4 months of Rifampicin (R), Isoniazid (H) [2RHZE/4RH].
Sputum Microscopy and Mycobacterial Culture.Microscopy was performed for D0 M5 and M6; culture was done at D0.At each visit, 3-5 mL of sputum were collected and underwent decontamination, digestion, and centrifugation for concentration.The pellet was used for microscopy using Auramine /Rhodamine staining (BBL™ Becton Dickinson, Sparks MD, USA) and uorescence microscope for reading the acid-fast bacillus (AFB).GeneXpert MTB/RIF® was performed to con rm the presence of Mycobacterium tuberculosis complex (MTBC) and determine rifampicin susceptibility.Mycobacteria culture was performed by inoculating both media, liquid Mycobacterium Growth Incubator Tube (MGIT), BD, USA, and solid (Middlebrook 7H11 agar and selective 7H11 agar).MTBC strains were identi ed by the Capilia method.Blood test.Each patient was tested for Human Immunode ciency Virus (HIV) using a discriminatory rapid test (SDBioline®).Patients with positive results were con rmed by ELISA.A random blood glucose test was performed to screen for diabetes and all high blood glucose levels were repeated for con rmation.Kato-Katz test for Schistosoma mansoni.One stool sample was obtained from each participant at D1 or during the rst week of the recruitment.The Kato-Katz test is a qualitative and semi-quantitative technique, recommended by the WHO for areas where the prevalence of schistosomiasis is between 10-50% [16].Data analysis.Data were entered into Excel 2013, cleaned, and analyzed by SPSS software version 25.0 for Windows.We performed descriptive analyses and compared means for age and body mass index (BMI).A comparison was done between TB/S.mansoni coinfected and TB mono-infected patients using independent sample T-tests.Binary and multiple logistic regressions were performed for associations between S. mansoni coinfection and sociodemographic and clinical characteristics.Any difference observed with a p-value < 5% at a 95% con dence interval (CI) was considered signi cant.

Discussion
Socio-Demographic Characteristics of Tuberculosis Patients.Males were predominant in our sample with 62.6%.The male tendency in active TB patients has been consistently reported by WHO and many studies [1,[17][18][19][20].The mean age was 34.9 ± 13.8 years 62.7% were years.In the African region and most high-burden countries in the world, TB is diagnosed in young people [1].In Mali, a study on TB in rural community settings found 79.2% of males [21].In Côte d'Ivoire, 70% of TB patients were under 35 years with a mean of 31 years [22].In the Central African Republic, where TB incidence reached 540/100,000 people, the mean age was 35.7 years and more than half were < 35 years [23].Occupations exposed to inhaled particles and promiscuity increase the risk of acquiring TB infection [24].Our study found a predominance of housewives, laborers, civil servants, and traders.In Mali and most endemic countries, TB treatment is provided on an outpatient basis, face mask-wearing is recommended for all patients, but not compulsory.Most of our patients were permanent residents of four administrative regions including Bamako (39.1%),Mopti (12.6%),Kayes (10.9%), and Ségou (9.2%), all located in the south alongside the two main rivers in Mali named Niger and Senegal.The Niger crosses the country over 1750 km, 42% of its total length, bathing Bamako, Koulikoro, Segou, Mopti, Timbuktu, Gao, and Ansongo [25,26].In our study, 55.2% of patients were living near a watercourse.These rivers serve for shing, rice cultivation, vegetable growing, and household work.In Mali, people in rural areas live in precarious hygiene due to poor socio-economic conditions.In 2020, the sanitation access rate was 39% [27].A community-based TB study found that farmers and housewives were the most dominant occupations with 58.3% and 20.8%, respectively [21].

Clinical Characteristics and Risk Factors for Reactivation of Latent Tuberculosis Infection.
The most known risk factors for LTBI reactivation are undernourishment, HIV/AIDS, diabetes, smoking, and alcoholism.In our study, undernourishment (BMI < 18.5 kg/m 2 ) was found in 55.7%, smoking (29.9%),HIV (11.5%), alcohol (13.8%), and diabetes (6.3%).Chest X-rays found bilateral pulmonary lesions in 51.1% and cavities in 41.2%.The bacteria load on microscopy was 3 + in 71.9% and 2 + in 22.4%.In the literature, the proportion of LTBI reactivation risk factors varies across studies.In Mali, the

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2023 (2.5 years).The study was conducted at the University Clinical Research Center (UCRC) of the University of Sciences, Techniques and Technologies of Bamako (USTTB) in Mali.Patients were enrolled and followed in 6 different TB diagnosis and treatment centers (DTC) and the Department of Pneumophtisiology of the University Teaching Hospital of Point-G, and the stools were tested at the clinical laboratory of Point-G Hospital.

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Table 1
Most patients were relatively young, male, married, with a primary or lower education level, informal occupation, and having their permanent residents near a river.Lung lesions were bilateral in 51.1% and cavity in 41.2%.Most patients, 71.9%, had positive sputum with 3 crosses on microscopy (Table

Table 2
Clinical characteristics of patients and Tuberculosis Risk Factors The proportion of known TB risk factors were smoking 30%, HIV 11.5%, undernourishment 55.7%; 71.9% had a high sputum bacteria load and 28.7% had Schistosoma mansoni coinfection.

Table 3
Outcome of patients followed during six (6) months of treatment.
Unfavorable treatment result was found in 11.3% of TB mono-infected versus 28% of TB/S mansoni co-infected patients.The death rate was 5.6% among TB mono-infected versus 18.0% among TB/S.mansoni co-infected patients.

Table 4
Factors associated with Schistosoma mansoni coinfection in tuberculosis patients.